Effects of nobiletin on intestinal stem cell proliferation <italic style="font-style: italic">in vitro</italic> and <italic style="font-style: italic">in vivo</italic>

Jingfeng WU; Kenan GUO; Ning ZHANG; Tianyu WU; Zimo ZHOU; Zhiyu LU; Fengchao WANG; Wensheng CHEN

doi:10.16016/j.2097-0927.202309138

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Effects of nobiletin on intestinal stem cell proliferation in vitro and in vivo

Basic Medicine | 更新时间：2026-01-09

- Effects of nobiletin on intestinal stem cell proliferation in vitro and in vivo
- Journal of Army Medical University Vol. 45, Issue 21, Pages: 2195-2205(2023)
- 作者机构：
  
  1.400038 重庆，陆军军医大学(第三军医大学)第一附属医院消化内科
  2.400038 重庆，陆军军医大学(第三军医大学)军事预防医学系复合伤研究所，创伤与化学中毒全国重点实验室
  3.400038 重庆，陆军军医大学(第三军医大学)基础医学院实验动物学教研室
- 作者简介：
- 基金信息：
  
  the General Program of National Natural Science Foundation of China(81872556);the Project of Science and Technology Innovation Capacity Promotion of Army Medical University(2021XJS03);the Chongqing Academician Program(基础研究与前沿探索类：cstc2018jcyj-yszxX0004)
- DOI：10.16016/j.2097-0927.202309138
  CLC： R285.5;R322.45;R329.28
- Received：25 September 2023，
  
  Revised：2023-10-13，
  
  Published：15 November 2023
- 稿件说明：
移动端阅览
Jingfeng WU, Kenan GUO, Ning ZHANG, et al. Effects of nobiletin on intestinal stem cell proliferation in vitro and in vivo[J]. Journal of Army Medical University, 2023, 45(21): 2195-2205.
DOI：

Jingfeng WU, Kenan GUO, Ning ZHANG, et al. Effects of nobiletin on intestinal stem cell proliferation in vitro and in vivo[J]. Journal of Army Medical University, 2023, 45(21): 2195-2205. DOI： 10.16016/j.2097-0927.202309138.

摘要

目的

探讨在体内外模型中常用有效剂量川陈皮素(nobiletin

NOB)对肠道干细胞的调节作用。

方法

构建小鼠结肠癌细胞系MC38的3D培养模型，加入不同浓度NOB，观察克隆死亡和存活。构建小鼠小肠类器官模型，在类器官培养体系中添加不同浓度的NOB，观察类器官的出芽和生长，利用MTT染色后的类器官成像进行面积和吸光值计算。在50、100 μmol/L浓度NOB干预体系中不同时间撤掉NOB，观察类器官生长。在50 μmol/L浓度NOB处理类器官的体系中分别加入R-spondin1、CHIR99021加强Wnt通路激活信号，观察类器官的生长。对C57/B6J小鼠灌胃不同浓度NOB，连续给药4 d，观察肠道隐窝和绒毛长度比，观察隐窝细胞凋亡，以及隐窝底部干细胞龛中Olfm4/BrdU染色阳性的细胞数量。

结果

与对照组相比，50 μmol/L NOB即可显著促进肿瘤细胞MC38成球后死亡，显著降低克隆形成率(

0.001)。与对照组相比，NOB从50 μmol/L起可以显著抑制肠类器官的出芽，50~200 μmol/L NOB可以显著抑制类器官的生长(

0.001)，并且具有剂量依赖效应；NOB撤药后在50 μmol/L时肠类器官

生长容易恢复，在100 μmol/L时难以恢复到正常水平。增强Wnt通路的激活可以部分挽救NOB对肠类器官的抑制作用。在体给予高于常用浓度的NOB不能诱导肠隐窝细胞凋亡，不影响肠干细胞的数目和增殖，不影响隐窝和绒毛比。

结论

NOB具有抑制正常肠干细胞的能力，可能通过调控干细胞相关通路如Wnt发挥作用，但其在在体不能发挥抑制干细胞增殖的效应；这提示NOB在常规浓度应用的安全性，同时提醒对NOB在在体肿瘤模型中发挥的抑制作用需要进行更深入的评估。

Abstract

Objective

To investigate the regulatory effect of nobiletin (NOB) at typical effective doses on intestinal stem cells

in vivo

and

in vitro

Methods

After a 3D culture model of mouse colorectal tumor cell line MC38 was constructed

the death and survival of the obtained colonies were observed after treatment of different concentrations of NOB. Mouse small intestinal crypts were cultured in the Matrigel to generate organoids. Different concentrations of NOB were added to the culture system

and the sprouting and growth of the organoids were observed. MTT assay was used to calculate the area and absorbance values of the organoids after staining. The organoid growth was observed by removing 50 and 100 μmol/L NOB for different periods. The effects of exogenous R-spondin1 and CHIR99021 (activators of Wnt pathway) on the growth of organoids were observed in the culture system with 50 and 100 μmol/L NOB treatment. C57/B6J mice were infused with different concentrations of NOB by gavage for 4 consecutive days. The ratio of intestinal crypt to villus length

cell apoptosis

and number of OLfm4 and BrdU double positive cells were calculated and observed.

Results

Compared with the control group

50 μmol/L NOB significantly promoted the death of MC38 cells after sphere formation and significantly reduced the colony formation rate (

0.001). The dose also significantly inhibited the budding of intestinal organoids

and 50~200 μmol/L NOB significantly inhibited the growth of intestinal organoids in a dose-dependent manner when compared with the contr

ol group (

0.001). After NOB withdrawal

the growth of intestinal organoids in the 50 μmol/L group was partially recovered

but it was difficult to recover to normal level in the 100 μmol/L NOB group. Enhanced Wnt pathway activation could partially rescue the inhibitory effect of NOB on intestinal organoids.

In vivo

administration of higher concentrations of NOB did not induce apoptosis of intestinal crypt cells

did not affect the crypt to villus ratio

and had no effects on the number and proliferation status of intestinal stem cells.

Conclusion

NOB possesses the ability to inhibit normal intestinal stem cells by regulating stem cell-related pathways such as Wnt

but this cellular inhibition cannot be phenocopied

in vivo

. Our study suggests the safety of NOB at regular concentration and also raises a caution to explain the inhibitory role of NOB in cancer models.

关键词

Keywords

references

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Effects of nobiletin on intestinal stem cell proliferation in vitro and in vivo

DOI：10.16016/j.2097-0927.202309138

摘要

Abstract

关键词

Keywords

references